This subproject is one of many research subprojects utilizing the resources provided by a Center grant funded by NIH/NCRR. Primary support for the subproject and the subproject's principal investigator may have been provided by other sources, including other NIH sources. The Total Cost listed for the subproject likely represents the estimated amount of Center infrastructure utilized by the subproject, not direct funding provided by the NCRR grant to the subproject or subproject staff. Objective: To advance human embryonic stem cells and their derivatives towards clinical application for treatment of blood disease. Understanding the mechanisms that regulate hematopoietic stem cell development is essential for further improvement of hematopoietic stem cell use in oncology and gene therapy. The overall goal of this project is to identify and characterize the hierarchy of progenitors and molecular pathways leading to hematopoietic stem cell development from human embryonic (hES) and induced pluripotent stem (iPS) cells. In our laboratory, we established a system for efficient hematopoietic differentiation of hES cells through coculture with OP9 bone marrow stromal cells. Using this system we were able to directly differentiate hES cells into cells of all major blood lineages (erythroid, myeloid and lymphoid), as well as identify different stages of hematopoietic commitment. We found that the earliest hematopoietic progenitors (HPs) in humans arise within CD34+ population and could be ultimately defined by surface expression of leukosialin (CD43). In addition, within CD43+ population, we identified lin-CD34+CD43+CD45- hematopoietic progenitors capable of differentiating toward all blood lineages including lymphoid cells, suggesting their hierarchical proximity to hematopoietic stem cells. However, molecular profiling of hES lin- CD34+CD43+CD45- cells revealed altered expression of genes associated with hematopoietic stem cell self-renewal and survival, reflecting limited engraftment potential of ES cell-derived hematopoietic progenitors. In addition we identified a novel precursor for mesenchymal stem and endothelial cells mesenchymonangioblast. With increasing interest in potential therapeutic application of hES cell derivates, identification of genes essential for hematopoietic stem cell development and diversification is of particular importance. The described experimental system sets a solid platform to advance in this direction. This research used WNPRC stem cell resources. PUBLICATION: Vodyanik MA, Yu J, Zhang X, Tian S, Stewart R, Thomson JA, Slukvin II. A mesoderm- derived precursor for mesenchymal stem and endothelial cells. Cell Stem Cell. 2010 Dec 3;7(6):718-29. PMID: 21112566, PMCID: PMC3033587.